New and improved – the next generation of GFP?

A new and improved green fluorescent protein, named mNeonGreen, was developed.

It was engineered from a Yellow fluorescent protein (LanYFP) that was isolated from the cephalochordate Branchiostoma lanceolatum. Therefore, LanYFP is genetically unrelated to the commonly used Aequorea victoria GFP.

Directed evolution to make it a monomer produced the new, monomeric protein, mNeonGreen. The ex/em of mNeonGreen is slightly shifted to the yellow compared to EGFP (509/516 compared to 488/509), making it a better choice to separate from CFP emission.

Though slightly less brighter than its parent protein LanYFP, mNeonGFP is 2-3 times brighter than EGFP and actually brighter than most green & yellow proteins.

Another great advantage of this new protein is that is it fast folding – the authors claim it is <10 min at 37C. This is fairly close to the superfolder GFP.

It is also very photostable (comparable to EGFP), performs well as a fusion construct at N & C termini many tested proteins, performs 4-times better that EGFP in stochastic single-molecule superresolution imaging and is a better FRET partner (both as acceptor and donor) than other proteins.

In short, this may very well be the “next generation” of fluorescent proteins. It has all the good qualities, and seems to have none of the bad ones. It performs better than most, if not all fluorescent proteins in every tested parameter.

Only its name is rather plain. I would call it something like wonderGFP or GreenLantern (hey, it even has the Lan from the animal they developed it from).

(Update: see here for details on how to get your hands on this protein).

I don’t know. We made stable human iPSCs expressing P2A-mNeonGreen and P2A-eGFP and eGFP was appreciably brighter. I was very hopeful we had a winner. Not sure why but our experience doesn’t really jive with the reportedly 3X brighter Neon signal. Possible cell type specific differences? Interference with P2A? Just don’t know. BTW – we used a standard GFP filter or FITC filters for detection and an XCITE broad spectrum light source for excitation.

That’s disapointing. I assume you checked there are no mutations in your construct. If it is not a cell-type of fusion protein issues, then I can only guess that maybe it is a matter of filters. Remember that mNeon is slightly yellow-shifted compared to GFP. Did you try a YFP filter?

The fate of the messenger is pre-determined

Meta

Disclaimer

The opinions expressed in this blog are solely those of the author. Any advice given in this blog is based on the author's knowledge and experience, which may be lacking. Therefore, the author of this blog may be wrong (don't tell his wife!).
Images are used here for research and criticism in accordance with fair use policies.